A cytokine is a greatly important proteinous substance as a biophylactic factor which closely relates to various kinds of antigen-specific, non-specific immune inflammatory responses. The existance thereof is necessary and indispensable for maintaining biological homeostasis and the cytokine is produced excessively in a disease with inflammation, relating to the formation and the prolongation of pathology of the disease.
"IL-8" is one kind of cytokine purified as a monocyte-derived neutrophil chemotactic factor (MDNCF) by Matsushima et al. and gene thereof has also been cloned in 1987. IL-8 is a neutrophile-activation migration regulatory factor produced by various kinds of cells.
It is known that IL-8 acts on a neutrophil to enhance functions for inhibiting growth of Candida albicans and Mycobacterium which the neutrophil has, and it is anticipated that IL-8 has an effect as an immunoactivation agent (Nipponigakukan, Cytokine--From Foundation to The Latest Information--, 1991, pages 65-72). Therefore, there has been desired a method for efficiently purifying IL-8 in a large amount from a body fluid or a culture medium.
On the other hand, the continuous administration of IL-8 in a large amount is remarkably harm to tissues, and causes the destruction of tissues of adult respiratory distress syndrome in an alveolus and the destruction of tissues with the infiltration of lymphocytes in a large amount in an arthrosis. Experimentally, it is shown that IL-8 relates essentially to the infiltration of neutrophils in dermatitis derived from lipopolysaccharide (LPS) and during reperfusion after ischemia. This has been proved by the fact that the destruction of tissues accompanied by the above-mentioned infiltration of lymphocytes or neutrophils can be inhibited almost completely by administrating a neutralizing antibody against IL-8. Further, an abnormally higher concentration of IL-8 has been detected in an inflammatory site or in peripheral blood of patients with diseases such as rheumatoid arthritis (RA), gouty arthritis, psoriasis, contact dermatitis, septicemia, idiopathic fibroid lung, adult respiratory distress syndrome, inflammatory bowel disease, immune angiitis, glomerular nephritis, urinary tract infection, cardiac infarction, asthma, respiratory tract infection, perinatal infectious disease and rejection in organ transplantation, than that of a normal human (refer to Menekiyakuri, 12, No. 1, pages 15-21 (1994)). And, it is considered that IL-8 relates to these diseases. At present, however, an efficient method to inhibit the function of IL-8 in these diseases has not been established.
Then, "IL-1.beta." is an inflammatory cytokine which is produced mainly by a monocyte and/or a macrophage with stimulation by a foreign matter such as bacteria, and in 1984, the gene of human has been cloned by Auron et al.
As seen from the fact that IL- 1.beta. has been discovered as an endogenious pyrogen (EP), a leukocytic endogenious mediator (LEM), a lymphocyte activation factor (LAF), a B cell-activating factor (BAF) and the like until the standardized name of interleukin 1 (IL-1) was determined as the same substance at The 2nd International Lymphokine Workshop in 1979, the bioactivity thereof is shown in various functions.
Though IL-1.beta., which is a main mediator of an inflammation, plays an important role in various kinds of reactions including homeostasis of a living body in the ordinary state, it has been clear that IL-1.beta. induces the destruction of tissues or the formation and/or deterioration of pathology in the inflammatory diseases when IL-1.beta. is produced excessively or conteniously for a long term in some mechanism (Biomedica, 9, 1993, pages 703-707). The relation of IL-1.beta. to each of pathological conditions such as toxic syndrome including septicemia, RA, Lyme disease, osteoporosis, Kawasaki disease, gout, glomerulonephritis, ectatic cardiomyopathy, endometritis, premature labor, granuloma, acute myelogenous leukemia, Alzheimer disease, Down syndrome, hepatic fibrosis, hepatoma, further, alcoholic hepatitis is strongly suggested (Nipponigakukan, Cytokine--From Foundation to The Latest Information--, 1991, pages 13-20 and pages 177-187), and a method of specific inhibition of IL-1.beta. is hoped for and researched enthusiastically.
Though, as the representative, an antiIL-1.beta. antibody, an antiIL-1.beta. receptor antibody, an IL-1 receptor antagonist (IL-1ra) and the like were developed (Igakunoayumi, 167, 1993, pages 432-435) and a part thereof was subjected to a clinical testing of septicemia as an objective disease, each did not give an effect to be expected and has not yet been put into practice.
Also, "IL-6" was originally isolated as a differentiating factor of a B cell and the structure of the gene thereof was determined by Hirano, Kishimoto et al. in 1986. It is recognized that IL-6 has a function as a main mediator of inflammation, for instance, to work as a proliferating factor of myelocytoma, and to derive an acute phase protein in liver.
It is shown by the production of a transgenic mouse that the abnormal production of IL-6 brings about a polyclonal activation of a B cell and causes plasmocytoma Also, in general, it has been observed that IL-6 in blood shows a high value in many acute inflammatory diseases or bacterial infection, virus infection, scald, myocardial infarction and the like. An example has been reported wherein IL-6 level was actually raised at the time of infestation such as scald or surgical operation and, in a cerebrospinal fluid of a patient with acute bacterial meningitis (by pneumococcus, staphylococcus, Listeria), up to 500 ng/ml of IL-6 was detected. On the other hand, IL-6 was detected in local inflammatory tissues or a general body of chronic hepatitis. In an autoimmune disease such as RA, Castleman disease or atrial myxoma, there is the abnormal production of IL-6 and it is considered that this triggers the production of proteins in hypergammaglobulinemia. In addition, it is suggested that the involvement of IL-6 to the diseases such as cancer of the uterine cervix, AIDS, alcoholic hepatitis, multiple myeloma, Lennert T lymphoma, mesangial proliferative nephritis, renal cytoma psoriasis and septicemia (Nipponigakukan, Cytokine--Foundation to The Latest Information--, 1991, pages 177-187). For the time being, however, an effective method for inhibiting specifically the function of IL-6 in these diseases has not yet been established.
Further, "IL-2" is a glycoprotein having a molecular weight of approximately 15 kDa which was conventionally called a T cell growth factor (TCGF) and is a cytokine of which gene was cloned by Taniguchi et al. in 1983. It is intended that the application of IL-2 as a remedy for cancer according to single therapy or local adoptive immunotherapy since IL-2 has a proliferation facilitating activity against T cell.
On the other hand, the opinion was obtained, suggesting that IL-2 continuously produced at a locality (in this case, at pancreas) is one of potent pathogenesis of autoimmune diabetes mellitus from an experiment using a transgenic mouse (Heath W. R., Allison J. et al.: Nature, 359, page 547, 1992). Also, it was also reported that the administration of IL-2 caused the development of both systemic lupus erythematosus (SLE) and RA (Chazerain P., Meyer O., Kahn M.-F., Ann. Intern. Med., 116, page 427, 1992 and Wandl U. B., Nagel-Hiemke M., May D. et al.: Clin. Immunol. Immunopathol., 65, page 70, 1992). Further, the possibility is suggested that IL-2 together with TNF-.alpha. relates closely to the pathologic conditions in the septic shock (Endo S., Inada K., Inoue Y. et al.: Circulatory Shock, 38, page 264, 1992). For the time being, however, an effective method for inhibiting the function of IL-2 in these diseases has not yet been established.
By considering the above-mentioned situation, the inventors of the present invention researched with respect to a method for adsorbing and removing IL-8, IL-1.beta., IL-6 and/or IL-2 as a pathogenic substance from a body fluid of a patient and, if necessary, a method for recovering the same. As the result of repeating whole-hertedly investigation, the inventors discovered that a water-insoluble carrier having an anionic functional group, when in contact with a body fluid, adsorbs strongly IL-8, IL-1.beta., IL-6 and IL-2 in the body fluid and proceeded the investigation to reach the completion of the present invention.
An object of the present invention is to selectively adsorb and remove IL-8, IL-1.beta., IL-6 and/or IL-2 from a body fluid, particularly, blood, plasma and serum and, if necessary, to recover the same.